The 5' untranslated leader region of HIV-1 and HIV-2 genomic RNA contains sequences essential for viral replication, including signals for transactivation of transcription, primer binding, splicing, encapsidation, RNA dimerization, and initiation of translation of the gag gene. Within a species, phylogenetic conservation of these non-coding sequences rivals even that of conserved coding domains for the viral proteins, attesting to their importance for replication. The high degree of conservation also highlights the potential to exploit this region for antiretroviral strategies. The use of conserved RNA sequences as drug targets has been well established, since many clinically useful antibiotics recognize a specific segment of bacterial rRNA. Biochemical characterization of the leader RNA of HIV-1 and HIV-2 suggests that it adopts multiple conformations that alternately display or hide RNA signals appropriate to different stages of the replication cycle. Based on the in vitro characterization of conformational isomers of HIV-2 leader RNA and upon the high degree of conservation of involved sequences, Dr. Lodmell hypothesizes that the leader region of HIV and other retroviruses is intimately involved in the regulation of several viral replicative processes, including dimerization of viral RNA, encapsidation, splicing, and protein synthesis, and that this regulation is manifested by differential presentation of the signaling structures found in the leader region. In this application, Dr. Lodmells proposes to test this hypothesis by constructing and characterizing viral mutants harboring substitution and deletion mutations designed to interfere with long- and short-range RNA interactions that were shown in vitro to be important in the modulation of RNA structure and behavior. A combination of cell culture, genetic, and biochemical approaches will be used to characterize the largely unexplored interrelationships of RNA splicing, dimerization, encapsidation, and translation in HIV types 1 and 2. The studies proposed here will aid in our overall understanding of the biochemistry and cell biology of human lentivirus replication as well as to validate the 5' leader region RNA as a potential antiretroviral target.